The possibility of obtaining the inclusion complex of γ-cyclodextrin nitrate with tetrahydrocarbazole and aminoadamantane conjugate, a promising drug for the treatment of neurodegenerative diseases, was investigated and proved using XRD, DSC, HPLC and NMR. It was found that the obtained complex, unlike the active substance itself, is soluble in water, and its cellular toxicity does not exceed that of the active substance, which contributes to the increase of its bioavailability and provides the possibility of conducting the necessary biological studies. It was found that the formed inclusion complex preserves the medicinal properties of the active substance, exhibiting mitoprotective and microtubule-stabilizing activities, and also possesses a pronounced antioxidant activity, effectively inhibiting spontaneous lipid peroxidation in rat brain homogenate.

Most of the medicinal products that are currently approved and used in clinical practice for neurodegenerative diseases, in particular Alzheimer’s disease, have a compensatory mechanism of action that enhances neurotransmitter signalling. It is an urgent need to develop new medicinal products combining cognitive-enhancing, neuroprotective, and disease-specific effects resulting from a multi-target mechanism of action including, in particular, prevention of glutamate-induced neuronal calcium uptake and stabilisation of microtubules.The aim of this study was to search for potentially neuroprotective and tauopathy-alleviating medicines amongst new thiouronium salt derivatives based on vicinal diamines.Materials and methods. The study investigated the ability of thiouronium salts to block glutamate-induced 45Ca2+ uptake by synaptosomes prepared from the brain of Wistar rats. The authors evaluated effects of these new compounds on polymerisation of a preparation of C57bl mouse brain tubulin and microtubule-associated proteins. The evaluation was carried out in the presence of guanosine triphosphate (GTP) and based on specific absorbance changes at 355 nm due to formation of microtubules. The authors analysed the structure of these microtubules, using negative staining followed by transmission electron microscopy. The IC50 determination and the statistical analysis were performed using standard software (Excel and PRISM 6.02).Results. The authors developed a screening algorithm for a number of new thiouronium salt derivatives based on vicinal diamines and studied biological activity of these derivatives by the effects on glutamate-induced calcium uptake by synaptosomes and on microtubule assembly processes. The authors identified compounds suppressing glutamate-induced calcium uptake by synaptosomes, i.e. compounds with neuroprotective potential. In addition, a number of new compounds were able to stimulate GTP-dependent microtubule assembly processes. The authors observed formation of microtubules with a normal structure in the presence of isopropyl-N’-[2-(benzoylamino)-1,2-diphenylethyl]-N-ethylimidothiocarbamate hydrobromide and considered the compound a promising scaffold for further optimisation.Conclusions. Chemical modification of thiouronium salts is a promising direction for developing effective neuroprotectors and microtubule stabilisers.

The conjugate of 2-methoxyestradiol, a ligand for the cell protein tubulin, with two molecules of chlorambucil, known as a DNA-alkylating agent, is capable of blocking the dynamics of tubulin-based microtubules at concentrations of 10–100 µmol L−1, by stabilizing them without changing their structure. Three analogs of this conjugate, which do not contain chloroethylamino and/or methoxy groups at C(2), were synthesized. At these concentrations, these analogs have another, inhibitory or combined, effect on the microtubule polymerization. Based on the molecular modeling results, the binding site of the initial conjugate in tubulin was proposed.

A series of novel imidazole-containing ligands and their organotin complexes were synthesized and characterized by NMR, IR, MALDI and elemental analysis. Redox behavior was studied by cyclic voltammetry (CV). Antioxidant properties were estimated in model reactions of single-electron reduction (CUPRAC-test), scavenging of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and O2−. radical anion, enzymatic oxidation of linoleic acid by lipoxygenase and Fe3+-induced lipid peroxidation of rat liver homogenates. It was found that ligands and complexes both possess radical scavenging activity of prolonged action type. Compounds exhibited notable antioxidant activity in lipid peroxidation. Cytotoxicity was estimated in standard MTT-test on multiple cell lines. Compounds demonstrated high toxicity on colon carcinoma and breast cancer cells and based on obtained data, lead compound was proposed. Additional assays were carried out for the lead compound, including regular MTT-test on cancer cells possessing various resistant mechanisms and modified MTT-test on tumor tissue samples, obtained from patients as well as apoptosis and cell cycle studies. All organotin complexes were also studied for their influence on tubulin polymerization. It was demonstrated that obtained compounds demonstrate unorthodox activity, promoting microtubules assembly rate instead of inhibiting it. Significant influence of compound 5 on G2/M phase of cell cycle is in accordance with influence on tubulin polymerization and lets us to mark synthesized compounds as mitotic poisons. The results open up the scopes for the search of novel antitumor agents for treatment of advanced forms of cancer.

Two series of organotin(IV) carboxylates based on phenolic antioxidants RCOOH and R(CH2)(2)COOH (R = 3,5-di-tert-butyl-4-hydroxyphenyl) and natural bile (cholanic) acids of formulae (RCOO)(2)SnMe2 (1); (RCOO)(2)SnBu2 (2); (R(CH2)(2)COO)(2)SnMe2 (3); (R(CH2)(2)COO)(2)SnBu2 (4); (cholate)SnPh3 (5); (deoxycholate)SnPh3 (6); (lithocholate)SnPh3 (7); (cholate)SnMe3 (8); (deoxycholate)SnMe3 (9), and (lithocholate)SnMe3 (10) were synthesized and characterized by H-1, C-13 NMR, IR and elemental analysis to study their antioxidant and cytotoxic potential. Compounds 1 and 2 were found by X-ray diffraction analysis to be monomers in the solid state; moreover, the carboxyl groups are coordinated bidentately through O atoms. The distorted octahedron geometry around the Sn center in the monocrystals of 1, 2 was revealed. The antioxidant activity of compounds as radical scavengers and reducing agents was evaluated in spectrophotometrical tests with stable radical DPPH, reduction of Cu2+ (CUPRAC method) and interaction with superoxide anion radical. Cytotoxicity in vitro of compounds has been estimated on human cancer (colon cancer HCT-116, lung carcinoma A549, breast cancer MCF-7, neuroblastoma SH-SY5Y) cell lines and normal fibroblasts cells. The IC50 values varied in 0.002-100 mu M range for compounds 1-4 depend on the alkyl substituents at Sn center and carboxylate ligand structure. The influence on Fe3+-induced lipid peroxidation, mitochondrial potential and mitochondrial permeability and tubulin assembly have been studied for the compounds 1 and 2. It was shown that the complexes 1 and 2 slightly depolarize the mitochondria but don't influence the calcium-induced mitochondrial permeability transition. Organotin complexes with cholic and deoxycholic acids as ligands were revealed to induce apoptosis in A549 cells. The percentage of apoptotic cells were 27.9 and 27.1% for triphenyltin complexes with cholic and deoxycholic derivatives, respectively. (C) 2019 Published by Elsevier B.V.

Ester of the tubulin ligand, 2-methoxyestradiol, with two molecules of DNA alkylating agent chlorambucil was proposed as a “triple” mutual prodrug for blocking rapid metabolism of the initial steroid. The target conjugate was synthesized via the Steglich esterification. The biotesting results demonstrate a very low cytotoxicity to cancer and conditionally normal cells (EC 50 ~100 μmol L −1 ), a weak inhibition of the cell growth (A549: 20%/48 h), no stimulation of apoptosis, and no influence on the microtubule morphology of human lung carcinoma A549 cells and the ability to increase the rate of microtubule assembling. The sharp distinction of these properties from those for the initial 2-methoxyestradiol and chlorambucil molecules indicates that the synthesized prodrug is stable in vitro and demonstrates an expediency of its further testing in vivo .

This study presents the synthesis of binary tetrohydro-γ-carbolines with ditriazol spacers of varying length, which exhibit anticholinesterase and antioxidant activity, as compared to the original Dimebon prototype. Anticholinesterase activity suggests the potential ability of the new compounds to block β-amyloid aggregation induced by anticholinesterase, making them promising candidates for further research preparations for the treatment of Alzheimer's disease. Particular attention should be paid to the conjugate with an intertriazol hexamethylene spacer, which can be regarded as the leading compound in this series.

Новые синтезированные бинарные конъюгаты тетрагидро-γ-карболинов, содержащие дитриазольные спейсеры разной длины, обладали выраженной антихолинэстеразной и антиоксидантной активностью, потенциальной способностью блокировать индуцируемую ацетилхолинэстеразой агрегацию бета-амилоида, в отличие от исходного прототипа Димебона. Всё это делает их перспективными кандидатами для дальнейшего исследования в качестве препаратов для лечения болезни Альцгеймера. Особого внимания заслуживает конъюгат с межтриазольным гексаметиленовым спейсером, который можно рассматривать как препарат — лидер в данном ряду соединений.

The synthesized new binary conjugates of tetrahydro-γ-carbolines, which contained ditriazole spacers of different length, exhibited considerable anticholinesterase and antioxidant activity as well as the potential ability to block the acetylcholinesterase-induced aggregation of β-amyloid in contrast to the original prototype Dimebon. This makes the compounds promising candidates for further investigation as drugs for the treatment of Alzheimer's disease. Special attention should be given to the conjugate containing the hexamethylene intertriazole spacer, which can be considered as a leader in this series of compounds.

The review and summarizes own and literature data about the role of mitochondria as the important target in the search for drugs for the treatment of neurodegenerative diseases. Aging is a major risk factor for sporadic forms of various neurodegenerative diseases, including Alzheimer′s disease. One of the most argued and currently accepted theories is the Mitochondrial Free Radical Theory of Aging. Mitochondrial hypotheses of the development of sporadic forms of neurodegenerative diseases particularly Alzheimer′s disease, are closely connected with it. Impairments of mitochondrial functions lead to a decrease in their ability to regulate calcium homeostasis in the cell and to a decrease in the threshold for the induction of mitochondrial permeability transition (MPT) pores. MPT inhibitors can be considered as a promising approach to the treatment of neurodegenerative diseases, since these drugs can not only exhibit the properties of neuroprotectors, but also can provide normalization of synaptic activity due to increased calcium capacity of mitochondria. The review presents data on the number of MPT inhibitors, including endogenous compounds melatonin and N-acetylserotonin, their bioisosteric analogue Dimebon and a number of other compounds. The use of mitochondria as a basis for the formation of screening strategy for the search for compounds for the treatment of neurodegenerative diseases is of particular interest – both as a test of their potential toxicity, and as a basis for the creation of metabolic stimulants and drugs with neuroprotective and cognitive-stimulating effect.

Phosphorylation of τ-protein, a component of microtubules in brain neurons, is a key pathomorphological sign of Alzheimer's disease. The development of this intracellular defect can be promoted by Al3+, Fe3+, and Zn2+ ions. The concentrations of these ions in the brain are considerably elevated in Alzheimer's disease. We performed a comparative study of phosphorylation of microtubular proteins of rat brain in the presence of Al3+, Fe3+, and Zn2+ ions in concentrations of 10-500 μM and microtubular proteins of brain of patients with Alzheimer's disease. The most likely candidate for the role of a factor that promotes hyperphosphorylation of τ-protein is Al3+ in concentrations of 250 and 500 μM.

The paper considers the influence of the structure of aminoadamantane–carbazole conjugates linked by five different spacers on their ability to inhibit acetylcholinesterase, butyrylcholinesterase (BChE), and carboxylesterase, the radical scavenging and mitoprotective activity, and tubulin polymerization. The nature of the spacer was found to be determinant for the appearance of anti-BChE and radical-scavenging activity. Aminoadamantane–carbazole conjugates linked by flexible spacers proved to be most effective in these assays. The influence of the conjugates on mitochondrial functions and tubulin polymerization largely depends on the structure of the carbazole moiety. A series of lead compounds promising for further study was identified as potential drugs for the treatment of neurodegenerative diseases, in particular, Alzheimer’s disease.

A synthetic approach to the vinylation and allylation of tetrahydro-γ-carbolines with alkane dimethyl sulfonates in the presence of sodium hydride was developed. The effect of the synthesized 5-vinyl- and 5-allyl-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indoles on acetylcholinesterase, butyrylcholinesterase, and carboxylesterase activity, mitochondrial functions, and tubulin polymerization was investigated, and their antioxidant activity was evaluated.

The article provides an overview of our own results of comparative study of influence of ions of iron, zinc and aluminium on the structure of microtubules from tubulin and microtubules associated proteins of rat brain with data on the structure of microtubules from tubulin and microtubules associated proteins from the brain of patients with Alzheimer′s disease (AD). A significant decrease in the amount of soluble tubulin was found in the postmortem brain of AD patients in comparison with the control group in the hippocampus, frontal cortex and substantia nigra, but not in the inferior olive. In vitro polymerization of tubulin and microtubules associated proteins from the brain of AD patients and electron micrographs of microtubules were obtained. The assembly of microtubules from brains of AD patients is disrupted, resulting in defective structures. On the other hand, the study of the influence of Al3+, Fe3+, Zn2+ on the microtubules from rat brains tubulin and microtubules associated proteins assembly and structure has shown that all studied metals are able to reduce the amount of microtubules and induce the assembly of anomal structures. According to the degree of the destructive effect on the microtubules and, accordingly, the possible significance in the pathogenesis of Alzheimer disease, metal ions can be arranged in the following sequence Al3+ > Zn2+ > Fe3+. Moreover, phosphorylation of tubulin and microtubules associated proteins in the presence of aluminum ions to the greatest extent reflects the phosphorylation of these proteins at AD. Comparison of data on the structure of microtubules after their assembly from brains of AD patients tubulin and microtubules associated proteins from brains of AD patients, and from the brain of rats, but in the presence of metal ions, confirm the conclusion about the possible role of the metals in the AD etiopathogenesis.

Novel gold(I) complexes, PPh3AuSR (1) and [(Ph3PAu)2SR]2(BF4)2 (2), with a protective antioxidant group R (R = 3,5-di-tert-butyl-4-hydroxyphenyl) were synthesized and characterized by 1H, 13C, 31P NMR, IR and elemental analyses. The crystal structures of compounds 1 and 2 were determined by X-ray diffraction. Compound 2 has a cluster structure with four Au–Au bonds in the crystal. The tetragold(I) dication is composed of four PPh3Au units with bridging sulfur atoms, where the Au4 moiety has a distorted tetrahedral geometry. The biological impact of gold complexes 1 and 2, and their precursors (RSH and AuPPh3Cl), on lipid peroxidation, mitochondrial functions, the tubulin polymerization, activity of glutathione reductase and cell viability were investigated. A comparative study of the structure–activity relationship was performed. We found that RSH and Au complexes 1 and 2 as opposed to AuPPh3Cl, prevented Fe3+ and tBHP-induced lipid peroxidation in isolated mitochondria. The number of 2,6-di-tert-butylphenol fragments influences the inhibitory effect. AuPPh3Cl induced swelling and depolarization of mitochondria, whereas RSH and the gold complexes 1 and 2, containing 2,6-di-tert-butylphenol groups, showed little to null effect. The thiol ligand RSH and AuPPh3Cl as well as gold complexes 1 and 2 have no impact on tubulin polymerization. The gold complexes were found to inhibit glutathione reductase. The compounds were tested for cytotoxicity against a primary culture of rat cerebellar granule cells. A high toxicity was determined for AuPPh3Cl, while introduction of antioxidant phenol groups decreased the cytotoxicity of the Au complexes. These results open up scope for the design of novel low toxic, gold based pharmacological agents.

Earlier unknown conjugates of carbazoles and γ-carbolines were obtained by the reaction of 3,6-substituted 9-oxiranylmethylcarbazoles and 2,8-substituted 2,3,4,5-tetrahydro-1H-pyrido-[4,3]indoles. The effects of synthesized 1-(9H-carbazol-9-yl)-3-(1,2,3,4-tetrahydro-5H-pyrido-[4,3-b]indol-5-yl)propan-2-ones and their hydrochlorides on neuronal NMDA receptors, polymerization of tubulin to form microtubules, and functional characteristics of rat liver mitochondria were studied.

Al(3+), Fe(3+), and Zn(2+) ions can disturb microtubule assembly from tubulin and microtubuleassociated proteins in rat brain. The main structural forms of these microtubules are rings and tangled bundles. These structures are formed only in the presence of Al(3+) and Fe(3+) ions. Therefore, Zn(2+) ions can be excluded from possible causes of structural abnormalities in microtubules during Alzheimer's disease. Al(3+) ions are the most probable etiological cause of Alzheimer's disease. The concentration of Al(3+) ions affecting the structure of microtubules is one order of magnitude lower than that of Fe(3+) ions (10 and 100 μM, respectively), which corresponds to their brain concentration reported in Alzheimer's disease.

Unknown conjugates of carbazoles and tetrahydrocarbazoles were obtained by the alkylation of carbazoles and tetrahydrocarbazoles with 3,6-substituted 9-oxiranylmethylcarbazoles. The influence of synthesized 1-(9Н-carbazol-9-yl)-3-(1,2,3,4-tetrahydro-5Н-pyrido[4,3-b]-indol-5-yl)propan-2-ones on functional characteristics of brain mitochondria has been studied. The potential neuroprotective activity of a several representatives of conjugates has been demonstrated using glutamate toxicity model.

We studied the effects of anti-Alzheimer drugs (tacrine, amiridine, and memantine) on phosphorylation of tubulin and microtubule-associated proteins isolated from rat brain, evaluated the capacity of these proteins to polymerize into microtubules after addition of study pharmacological agents, and analyzed the structure of generated microtubules. It was shown that test substances impair assembly of microtubules to a different extent. Dose-dependent effects of these agents on phosphorylation of tubulin and microtubule-associated proteins were observed. Triazolam (not approved for clinical use as anti-Alzheimer drug) in the same concentrations was used as the reference substance in the same tests. It was observed that this substance even in minimal concentration induced the most pronounced changes in microtubule structure. A direct correlation between the capacity of the test substances to modulate tubulin phosphorylation and to impair microtubule structure was found: the more the substance inhibited tubulin phosphorylation, the more it disordered microtubule structure.